Chordomas are rare, slow-growing, malignant sarcomas that originate from notochordal remnants along the axial skeleton. Molecular mechanism of chordoma development remains unknown. Intriguingly, tuberous sclerosis complex (TSC) is the only known syndrome where the incidence of chordomas has been described. TSC is an autosomal dominant tumor suppressor syndrome characterized by abnormal tissue growths, known as hamartomas, in several organs. In an earlier study from our laboratory, biallelic mutations of TSC genes were identified for the first time in chordomas obtained from TSC patients. TSC proteins tuberin and hamartin function together to inhibit mammalian target of rapamycin (mTOR) pathway, a key regulator of protein synthesis and cell growth. In addition to TSC genes, other tumor suppressor genes such as PTEN, NF1 and LKB1 critically regulate mTOR signaling through TSC proteins. Based on these, we hypothesize that mutations in tumor suppressors, which negatively regulate mTOR signaling, may be associated with sporadic chordoma pathogenesis. Our preliminary data demonstrate that sacral chordoma tumors and chordoma-derived cell lines commonly display constitutively activated mTOR signaling and deficiency of PTEN expression. These suggest that loss of PTEN as well as other genetic alterations resulting in aberrant activation of mTOR signaling may contribute to the development of sporadic chordomas. In this proposal, we aim to examine mutations in candidate genes, including PTEN, TSC1, and TSC2, by direct sequencing and loss of heterozygosity (LOH) assays in 30 sporadic chordomas. Furthermore, we propose to generate two chordoma mouse models by deleting Tsc1 or Pten in the notochord using inducible Cre/loxP system. Currently, there is no valid animal model, which is a major obstacle for chordoma research and therapeutic development. If successfully carried out, this proposal would not only break new ground for understanding the pathogenesis of chordomas, but also provide the scientific community with valuable reagents, which can serve as model systems for drug screening and preclinical studies of chordoma. In addition, this study would provide a rationale for clinical trials of Akt/mTOR inhibition in patients with sporadic chordomas. PUBLIC HEALTH RELEVANCE: Chordoma is a slow-growing, malignant tumor, which usually occurs in the spine and base of skull. This project is aimed to examine genetic mutation in chordomas and to generate chordoma animal models, and thus has significance in public health.